Since the original introduction by Murphy and Riley of their method for determination of inorganic phosphorus, a great deal of interest in its use for the determination of phosphate in natural waters or body fluids such as blood serum has been expressed by researchers and commercial interests. Indeed, this and closely related methods are the only ones in common use for the colorimetric determination of phosphorus.
The Murphy and Riley method is based on the formation of a complex between phosphate and molybdate which can be quantitated by using its absorption peak in the ultraviolet range or by reducing the complex to produce the blue color of "molybdenum blue" and/or of a series of fairly complex structures known as "heteropoly blues". A significant part of the challenge of this work is to achieve a testing methodology having the necessary convenience of an on-site or field type device coupled with the required sensitivity, rapid rate of reaction and stable color suitable for quantitative work as well as to eliminate positive interferences that can be seen in the presence of silicates, arsenates, or proteins or negative interference from fluoride.
Over the years, researchers have devoted considerable effort to the solution of the above problems and to developing an understanding of the complexity of the chemical structures involved in the presentation of color. "Molybdenum blue" is now understood to have a large structure involving as many as 154 molybdenum atoms, and reaction conditions are thought to determine the combination of "heteropoly blues" that are formed. Because of the complexity of interferences observed, various reactions and associated rates, and incompletely characterized product mixtures, workers have traditionally struggled to achieve reliability and reproducibility in tests based on quantitation of this blue color.
As an exemplary illustration of the application of the present invention, the management of soil chemistry made possible by soil testing is known to farmers to play an important part in the successful production of fruits and vegetables. Farmers typically sample various portions of their fields and send samples to professional laboratories for evaluation of the levels of chemical components including that of soil phosphorus. The Bray method is most commonly used for this and involves extraction of phosphate from the soil using an aqueous sodium bisulfate/sodium fluoride solution followed by reaction of the extract with molybdate and reduction of the resulting complex with ascorbic acid to form a blue color. The visible absorption peak is then used for quantitation in a spectrophotometer. Of course, the same need for testing applies to home gardeners of fruits, vegetables, or flowers, so there is a need for an inexpensive, reliable test method which can be used on site to rapidly obtain a useful result. Soil test kits currently available in this market generally require a cumbersome number of steps and relatively lengthy waiting periods between such steps.
The present invention describes a new way of measuring phosphorus concentration that has accuracy sufficient for the purpose and is faster and more convenient than previously known methods.